The present invention pertains to the discovery, isolation, characterization and utilization of a novel strain of porcine reproductive and respiratory syndrome (PRRS) virus. The invention further pertains to diagnostic and protective antigens and vaccines for the PRRS disease in pigs, and the methods of making and using the same.
All publications and patent applications herein are incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.
Porcine reproductive and respiratory syndrome (PRRS) was first described in the United States in 1987 and in Germany in 1990. Other countries subsequently reporting the disease include France (1), Denmark (2), the Netherlands (3), Japan (4), Canada (5), Spain (6), and England (7). PRRS has the potential to become an economic disaster for U S. and European swine producers. There is evidence of extreme genetic and antigenic variability between American and European isolates (29).
This syndrome has variously been called mystery swine disease, swine infertility and respiratory syndrome (SIRS), abortus blau, blue eared pig disease, xe2x80x98Lelystad virusxe2x80x99,xe2x80x9cHeko-hekoxe2x80x9d disease, and porcine epidemic abortion and respiratory syndrome (PEARS)(8-10).
The disease caused by the PRRS virus is characterized by: severe reproductive failure in sows, resulting in late term abortions with an inceased incidence of mummified, stillborn and weak pigs; chronic problems with delayed return to estrus; mild to severe respiratory distress, including sneezing, coughing, and nasal or ocular discharge in young pigs; increased mortality in preweaning, weaning and growing pigs; a mild flu-like disease in grower-finisher pigs; conjunctivitis, and lymph node enlargement (11-13). The respiratory syndrome is often associated with severe infection with secondary bacterial agents including Pasteurella multocida, Haemophilus parasuis and Streptococcus suis (8). Experimentally inoculated piglets showed clinical signs of depression, anorexia, pyrexia, diarrhea, sitting posture and periocular edema (12). One study estimated that the economic loss from PRRS on 91 Dutch farms averaged 29.5 kg per sow per year due to a decreased number of piglets raised per sow per litter, a prolonged farrowing interval and a higher replacement rate of sows (3).
The virus that is responsible for the disease is an enveloped RNA virus belonging to the Arterivirus group within the Togaviridae family. While data concerning morphology, morphogenesis and virion composition first suggested that the PRRS virus belonged to the Arterivirus group, this conclusion has been confirmed by analysis of genome organization, gene expression strategy and by comparison of deduced protein sequences. The arteriviruses also include the equine arteritis virus (EAV), lactate dehydrogenase elevating virus(LDV)and simian hemorrhagic fever virus (SHFV). The PRRS viruses are single stranded RNA viruses having a viral genome of positive polarity and a size of approximately 15 kb (15). These spherical virions are estimated to be 48-83 nm in diameter comprised of a 25-30 nm core surrounded by an envelope (3). The positive-strand RNA genome possesses at least three major structural proteins designated N, M, and E. The N protein is considered as the major component of the nucleocapsid, whereas M and E are membrane-associated (16). The nucleotide sequence of the genomic RNA of the PRRS virus has been determined for isolate CDI-NL-2.91 (Institut Pasteur deposit number I-1102) from overlapping cDNA clones (17). Eight open reading frames (ORFs) were identified in the consecutive sequence of 15,088 bp obtained for this isolate. The 3xe2x80x2-portion of the genome of another U.S. isolate, ATCCVR 2332, was also cloned and sequenced (18). The resultant 3,358 nucleotides contained 6 ORFs with homologies to ORFs 2 through 7 of the European strain of the PRRS virus and other members of the free-standing genus of arteriviruses.
As noted previously, there is evidence of extreme genetic and antigenic variability between different isolates of the PRRS virus. Thus, there is a need for the isolation, characterization, and utilization of novel PRRS viral isolates. Herein, we report on the discovery, isolation and use of the novel PRRS virus strain designated JK-100.
This invention comprises the identification and isolation of PRRS virus strains from selected piglets in infected herds, where the selected piglets showed signs of viremia and seroconversion but no clinical symptoms. More specifically, we have identified and isolated a novel PRRS viral strain designated JK-100.
The PRRS viral strain designated JK-100 has been deposited in accordance with the Budapest Treaty on the International Recognition of the Deposit of Microorganisms for the purpose of Patent Procedure with the China Center for Type Culture Collection, Wuhun University, Wuhun 430072, PRC, under the Designation CCTCC V 200005. The date of the deposit was Aug. 8, 2000.
The present invention also provides PRRS viral strains essentially corresponding to strain JK-100. The words xe2x80x9cessentially correspondingxe2x80x9d refer to variations that occur in nature and to artificial variations of JK-100, particularly those which still allow detection by techniques like hybridization, PCR and ELISA, using JK-100-specific materials, such as JK-100-specific antibodies.
The present invention provides for propagation of strain JK-100 to high titer in certain cell lines such as the MARC 145 cell line. This invention provides compositions of matter comprising the JK-100 strain. More specifically, this invention provides compositions of matter which comprise live, killed or attenuated JK-100 virus.
The present invention further provides a vaccine composition for vaccinating animals, in particular mammals, more particular pigs or swine, to protect them against PRRS disease. More specifically, the vaccine composition comprises the JK-100 viral strain, either live, killed, or attenuated; or an antigenic part or component of JK-100; a protein or antigenic polypeptide derived from, or a peptide mimicking an antigenic component of, JK-100; and a suitable carrier or adjuvant. Thus, the present invention also provides processes for preparing and using live virus or killed virus antigens (conventional or recombinant) and the vaccines resulting therefrom by combining an immunologically effective amount of the virus with diluent and/or adjuvant, respectively.
The invention also provides a diagnostic kit for detecting antigen from JK-100 in a sample, in particular a biological sample such as blood or blood serum, sputum, saliva, semen, or tissue, derived from an animal, in particular a mammal, more in particular a pig or swine, comprising an antibody which specifically recognizes a part or component of JK-100, and suitable detection means of an antigen detection assay. The invention further provides a diagnostic kit for detecting an antibody which specifically recognizes JK-100 in a sample, in particular a biological sample such as blood or blood serum, sputum, saliva, semen, or tissue, derived from an animal, in particular a mammal, more in particular a pig or swine, comprising JK-100; an antigenic part or component of JK-100; a protein or antigenic polypeptide derived from JK-100; or antigenic polypeptide derived from JK-100; or a peptide mimicking an antigenic component of JK-100; and suitable detection means of an antibody detection assay.
One skilled in the art can easily make any necessary adjustments in accordance with the necessities of the particular situation.